Calcineurin Aα but Not Aβ Augments ICl(Ca) in Rabbit Pulmonary Artery Smooth Muscle Cells

Abstract Activation of Ca2+-dependent Cl- currents (ICl(Ca)) increases membrane excitability in vascular smooth muscle cells. Previous studies showed that Ca2+-dependent phosphorylation suppresses ICl(Ca) in pulmonary artery myocytes, and the aim of the present study was to determine the role of the Ca2+-dependent phosphatase calcineurin on chloride channel activity. Immunocytochemical and Western blot studies with isoform-specific antibodies revealed that the α and β forms of the CaN catalytic subunit are expressed in PA cells but that only the α variant translocated to the cell periphery upon a rise in intracellular [Ca2+]. ICl(Ca) evoked by pipette solutions containing a [Ca2+] set at 500 nm was considerably larger when the pipette solution included constitutively active CaN containing the α catalytic isoform. This stimulatory effect was lost by boiling the enzyme or by the inclusion of a specific CaN inhibitory peptide and was not shared by the inclusion of the β form of the catalytic subunit. In the absence of constitutively active CaN, cyclosporin A, an inhibitor of CaN, suppressed ICl(Ca) evoked by 500 nm Ca2+ when the current amplitude was relatively large but was ineffective in cells with smaller currents. In perforated patch recordings, cyclosporin A consistently inhibited ICl(Ca) evoked as a consequence of Ca2+ influx through voltage-dependent calcium channels. These novel data show that in PA myocytes activation of ICl(Ca) is enhanced by Ca2+-dependent dephosphorylation and that the regulation of this conductance is highly isoform-specific.